Evaluation of a 99mTc-Labeled AnnexinA5 Variant for Non-invasive SPECT Imaging of Cell Death in Liver, Spleen and Prostate
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We investigate radio-labeling and pharmacokinetics of a new AnnexinA5 variant (HYNIC-cys-AnxA5) and then assess its utility for the non-invasive detection of cell death in liver, spleen and prostate.
AnnexinA5 binds to phosphatidylserine expressed on the surface of apoptotic and necrotic cells. Contrary to other AnnexinA5 variants, the new cys-AnxA5 allows for site-specific conjugation of a hydrazinonicotinamide-maleimide moiety and subsequent radio-labeling with 99mTc at a position not involved in the AnxA5-phosphatidylserine interaction. Distribution of 99mTc-HYNIC-cys-AnxA5 was studied in rats, both invasively and via SPECT/CT. Cycloheximide was used to induce cell death in liver and spleen, whereas apoptosis in the prostate was induced by castration.
HYNIC-cys-AnxA5 was efficiently and reproducibly labeled with 99mTc. Blood clearance of radioactivity after iv-injection was adequately described by a two-compartment model, the renal cortex representing the main site of accumulation. Cycloheximide treatment resulted in increased accumulation of intravenous-injected 99mTc-HYNIC-cys-AnxA5 in liver and spleen over controls, which correlated well with TUNEL staining for cell death in corresponding tissue sections. However, the increase in TUNEL-positive prostate epithelial cells observed following castration was not paralleled by greater 99mTc-HYNIC-cys-AnxA5 accumulation.
99mTc-HYNIC-cys-AnxA5 appears a suitable tracer for assessment of cell death in liver and spleen, but not prostate.
KEY WORDSannexinA5 apoptosis cell death molecular imaging SPECT/CT
AnnexinA5 with cysteine incorporated via site-directed mutagenesis
Region of interest
Standard Error of the Mean
Single Photon Emission Computed Tomography/Computed Tomography
Terminal deoxynucleotidyl transferase dUTP Nick End Labeling
Percentage of Injected Dose
C. van Kammen and C. van Helvert of the University of Maastricht are gratefully acknowledged for animal handling and experimentation, whereas R. van Brakel, M. Berben and S. Kivits are thanked for skillful conduction of the radio-labeling, biodistribution studies and SPECT/CT experiments, respectively. Finally, Drs. B. Henry, O. Steinbach, R. Rossin, C. P. Reutelingsperger, I. Verel, D. Attia, H. Hamersma and S. Addo are thanked for valuable scientific discussions.
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